Measurement of Reactive Oxygen Species and Nitric Oxide from Tomato Plants in Response to Abiotic and Biotic Stresses.

Nitric oxide (NO) is a free radical molecule that has been known to influence several cellular processes such as plant growth, development, and stress responses. NO together with reactive oxygen species (ROS) play a role in signaling process. Due to extremely low half-life of these radicals in cellular environment, it is often difficult to precisely monitor them. Each method has some advantages and disadvantages; hence, it is important to measure using multiple methods. To interpret the role of each signaling molecule in numerous biological processes, sensitive and focused methods must be used. In addition to this complexity, these Reactive Oxygen Species (ROS) and NO react with each other leads to nitro-oxidative stress in plants. Using tomato as a model system here, we demonstrate stepwise protocols for measurement of NO by chemiluminescence, DAF fluorescence, nitrosative stress by western blot, and ROS measurement by NBT and DAB under stress conditions such as osmotic stress and Botrytis infection. While describing methods, we also emphasized on benefits, drawbacks, and broader applications of these methods.

Mitochondrial alternative oxidase pathway helps in nitrooxidative stress tolerance in germinating chickpea.

Mitochondrial alternative oxidase (AOX) is an important protein that can help in regulating reactive oxygen species and nitric oxide in plants. The role of AOX in regulation of nitro-oxidative stress in chickpea is not known. Using germinating chickpea as a model system, we investigated the role of AOX in nitro-oxidative stress tolerance. NaCl treatment was used as an inducer of nitro-oxidative stress. Treatment of germinating seeds with 150 mM NaCl led to reduced germination and radicle growth. The AOX inhibitor SHAM caused further inhibition of germination, and the AOX inducer pyruvate improved growth of the radicle under NaCl stress. Isolated mitochondria from germinated seeds under salt stress not only increased AOX capacity but also enhanced AOX protein expression. Measurement of superoxide levels revealed that AOX inhibition by SHAM can enhance superoxide levels, whereas the AOX inducer pyruvate reduced superoxide levels. Measurement of NO by gas phase chemiluminescence revealed enhanced NO generation in response to NaCl treatment. Upon NaCl treatment there was enhanced tyrosine nitration, which is an indicator of nitrosative stress response. Taken together, our results revealed that AOX induced under salinity stress in germinating chickpea can help in mitigating nitro-oxidative stress, thereby improving germination.

Nitric oxide, energy, and redox-dependent responses to hypoxia.

Hypoxia occurs when oxygen levels fall below the levels required for mitochondria to support respiration. Regulated hypoxia is associated with quiescence, particularly in storage organs (seeds) and stem cell niches. In contrast, environmentally induced hypoxia poses significant challenges for metabolically active cells that are adapted to aerobic respiration. The perception of oxygen availability through cysteine oxidases, which function as oxygen-sensing enzymes in plants that control the N-degron pathway, and the regulation of hypoxia-responsive genes and processes is essential to survival. Functioning together with reactive oxygen species (ROS), particularly hydrogen peroxide (H2O2) and reactive nitrogen species (RNS), such as nitric oxide (·NO), nitrogen dioxide (·NO2), S-nitrosothiols (SNOs), and peroxynitrite (ONOO-), hypoxia signaling pathways trigger anatomical adaptations such as formation of aerenchyma, mobilization of sugar reserves for anaerobic germination, formation of aerial adventitious roots, and the hyponastic response. NO and H2O2 participate in local and systemic signaling pathways that facilitate acclimation to changing energetic requirements, controlling glycolytic fermentation, the γ-aminobutyric acid (GABA) shunt, and amino acid synthesis. NO enhances antioxidant capacity and contributes to the recycling of redox equivalents in energy metabolism through the phytoglobin (Pgb)-NO cycle. Here, we summarize current knowledge of the central role of NO and redox regulation in adaptive responses that prevent hypoxia-induced death in challenging conditions such as flooding.

Beneficial effects of bio-fabricated selenium nanoparticles as seed nanopriming agent on seed germination in rice (Oryza sativa L.).

Climate change and increasing population pressure have put the agriculture sector in an arduous situation. With increasing demand for agricultural production overuse of inputs have accentuated the negative impact on environment. Hence, sustainable agriculture is gaining prominence in recent times with an emphasis on judicious and optimum use of resources. The field of nanotechnology can immensely help in achieving sustainability in agriculture at various levels. Use of nutrients and plant protection chemicals in nano-form can increase their efficacy even at reduced doses thus decreasing their pernicious impact. Seed priming is one of the important agronomic practices with widely reported positive impacts on germination, seedling growth and pathogen resistance. In the current study, the effect and efficacy of selenium nanoparticles synthesized using phyto-extracts as a seed priming agent is studied. This nanopriming enhanced the germination, hastened the seedling emergence and growth with an increase in seedling vigour and nutrient status. This eco-friendly and economical method of synthesizing nanoparticles of various nutrient minerals can optimize the resource use thus helping in sustainable agriculture by reducing environment damage without compromising on efficacy.